title: Isofagomine induced stabilization of glucocerebrosidase.
abstract: Structurally destabilizing mutations in acid beta-glucosidase (GCase) can result in Gaucher disease (GD). The iminosugar isofagomine (IFG), a competitive inhibitor and a potential pharmacological chaperone of GCase, is currently undergoing clinical evaluation for the treatment of GD. An X-ray crystallographic study of the GCase-IFG complex revealed a hydrogen bonding network between IFG and certain active site residues. It was suggested that this network may translate into greater global stability. Here it is demonstrated that IFG does increase the global stability of wild-type GCase, shifting its melting curve by approximately 15 degrees C and that it enhances mutant GCase activity in pre-treated N370S/N370S and F213I/L444P patient fibroblasts. Additionally, amide hydrogen/deuterium exchange mass spectroscopy (H/D-Ex) was employed to identify regions within GCase that undergo stabilization upon IFG-binding. H/D-Ex data indicate that the binding of IFG not only restricts the local protein dynamics of the active site, but also propagates this effect into surrounding regions.
authors.affiliation: ExSAR Corporation, 11 Deer Park Drive, Suite 103, Monmouth Junction, NJ 08852, USA.
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Kornhaber
authors.forename: Michael B
authors.initials: MB
authors.lastname: Tropak
authors.forename: Gustavo H
authors.initials: GH
authors.lastname: Maegawa
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Tuske
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Coales
authors.forename: Don J
authors.initials: DJ
authors.lastname: Mahuran
authors.forename: Yoshitomo
authors.initials: Y
authors.lastname: Hamuro
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain; D002460:Cell Line; D041961:Deuterium Exchange Measurement; D004795:Enzyme Stability / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D005470:Fluorometry; D005962:Glucosylceramidase / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D050112:Imino Pyranoses / Q000378:metabolism / Q000494:pharmacology*; D008247:Lysosomes / Q000187:drug effects / Q000201:enzymology; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation
chemical_list: D050112:Imino Pyranoses; C098432:isofagomine; D005962:Glucosylceramidase